Abstract：
BACKGROUND: Jitai tablet, a traditional Chinese medicine, has a neuroprotective effect on 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine (MPTP)-induced Parkinson\'s disease (PD) mice. As one of the main active ingredients in the Jitai tablet, corydaline (Cory) has analgesic and anti-allergic effects, but it has not been studied in PD. Here, we investigated the role and mechanism of Cory in PD.
METHODS: The PD model was induced by MPTP. Cell viability was measured by 3-(4, 5)-dimethylthiahiazo (-z-y1)-3, 5-di-phenytetrazoliumromide assay. The Pole test and traction test were performed to detect the behaviors of mice. The expression of tyrosine hydroxylase (Th) was detected by immunohistochemistry and Western blot. Immunofluorescence staining, monodansylcadaverine staining, and Western blot were conducted to assess autophagy. A lactic dehydrogenase release assay was used to detect cytotoxicity. Network pharmacology was used to screen the targets.
RESULTS: There existed cytotoxicity when the concentration of Cory reached 40 μg/mL. Cory (not exceeding 20 μg/mL) could alleviate MPTP-induced cell damage. In vivo experiments indicated that Cory could improve the motor coordination of mice with PD. Besides, Cory could increase LC3-II/LC3-I levels both in vivo and in vitro. In addition, the Th levels reduced in the striatum and middle brain tissues of Parkinson\'s mice were recovered by Cory injection. We also found that Cory decreased the phosphorylation of glucogen synthase kinase-3 beta (GSK-3β) at Tyr216 and increased the phosphorylation of GSK-3β at Ser9 not only in primary neurons and SH-SY5Y cells but also in the striatum and middle brain tissues. Furthermore, Cory increased LC3-II/LC3-I levels and decreased p62 levels by regulating GSK-3β.
CONCLUSIONS: Cory enhanced autophagy, attenuated MPTP-induced cytotoxicity, and alleviated PD partly through the regulation of GSK-3β phosphorylation.
METHODS: The PD model was induced by MPTP. Cell viability was measured by 3-(4, 5)-dimethylthiahiazo (-z-y1)-3, 5-di-phenytetrazoliumromide assay. The Pole test and traction test were performed to detect the behaviors of mice. The expression of tyrosine hydroxylase (Th) was detected by immunohistochemistry and Western blot. Immunofluorescence staining, monodansylcadaverine staining, and Western blot were conducted to assess autophagy. A lactic dehydrogenase release assay was used to detect cytotoxicity. Network pharmacology was used to screen the targets.
RESULTS: There existed cytotoxicity when the concentration of Cory reached 40 μg/mL. Cory (not exceeding 20 μg/mL) could alleviate MPTP-induced cell damage. In vivo experiments indicated that Cory could improve the motor coordination of mice with PD. Besides, Cory could increase LC3-II/LC3-I levels both in vivo and in vitro. In addition, the Th levels reduced in the striatum and middle brain tissues of Parkinson\'s mice were recovered by Cory injection. We also found that Cory decreased the phosphorylation of glucogen synthase kinase-3 beta (GSK-3β) at Tyr216 and increased the phosphorylation of GSK-3β at Ser9 not only in primary neurons and SH-SY5Y cells but also in the striatum and middle brain tissues. Furthermore, Cory increased LC3-II/LC3-I levels and decreased p62 levels by regulating GSK-3β.
CONCLUSIONS: Cory enhanced autophagy, attenuated MPTP-induced cytotoxicity, and alleviated PD partly through the regulation of GSK-3β phosphorylation.
摘要：
背景：吉泰平板电脑，中药，对1-甲基-4-苯基-1，2，3，6-四氢吡啶（MPTP）诱导的帕金森病（PD）小鼠具有神经保护作用。作为吉泰片中的主要活性成分之一，Corydaline（Cory）具有镇痛和抗过敏作用，但尚未在PD中研究过。这里,我们研究了Cory在PD中的作用和机制。
方法：用MPTP诱导PD模型。细胞活力通过3-(4,5)-二甲基硫代偶氮(-z-y1)-3,5-二苯基四唑三虫测定来测量。进行了极点测试和牵引测试以检测小鼠的行为。免疫组化和Westernblot检测酪氨酸羟化酶(Th)的表达。免疫荧光染色,单丹西尸胺染色，并进行Westernblot以评估自噬。乳酸脱氢酶释放测定法用于检测细胞毒性。网络药理学用于筛选目标。
结果：当Cory浓度达到40μg/mL时，存在细胞毒性。Cory(不超过20μg/mL)可以减轻MPTP诱导的细胞损伤。体内实验表明，Cory可以改善PD小鼠的运动协调能力。此外,Cory可以在体内和体外增加LC3-II/LC3-I水平。此外,通过Cory注射恢复帕金森病小鼠纹状体和中脑组织中Th水平降低。我们还发现，Cory不仅在原代神经元和SH-SY5Y细胞中，而且在纹状体和中脑组织中，在Tyr216上降低了葡糖原合酶激酶3β（GSK-3β）的磷酸化，并在Ser9上增加了GSK-3β的磷酸化。此外，Cory通过调节GSK-3β增加LC3-II/LC3-I水平并降低p62水平。
结论：Cory增强自噬，减弱MPTP诱导的细胞毒性，并通过调节GSK-3β磷酸化部分减轻PD。
方法：用MPTP诱导PD模型。细胞活力通过3-(4,5)-二甲基硫代偶氮(-z-y1)-3,5-二苯基四唑三虫测定来测量。进行了极点测试和牵引测试以检测小鼠的行为。免疫组化和Westernblot检测酪氨酸羟化酶(Th)的表达。免疫荧光染色,单丹西尸胺染色，并进行Westernblot以评估自噬。乳酸脱氢酶释放测定法用于检测细胞毒性。网络药理学用于筛选目标。
结果：当Cory浓度达到40μg/mL时，存在细胞毒性。Cory(不超过20μg/mL)可以减轻MPTP诱导的细胞损伤。体内实验表明，Cory可以改善PD小鼠的运动协调能力。此外,Cory可以在体内和体外增加LC3-II/LC3-I水平。此外,通过Cory注射恢复帕金森病小鼠纹状体和中脑组织中Th水平降低。我们还发现，Cory不仅在原代神经元和SH-SY5Y细胞中，而且在纹状体和中脑组织中，在Tyr216上降低了葡糖原合酶激酶3β（GSK-3β）的磷酸化，并在Ser9上增加了GSK-3β的磷酸化。此外，Cory通过调节GSK-3β增加LC3-II/LC3-I水平并降低p62水平。
结论：Cory增强自噬，减弱MPTP诱导的细胞毒性，并通过调节GSK-3β磷酸化部分减轻PD。
